ALTERATIONS OF COPY NUMBER OF METHYLATION
PATTERN IN MISMATCH REPAIR GENES BY METHYLATION
SPECIFIC-MULTIPLEX LIGATION-DEPENDENT PROBE
AMPLIFICATION IN CASES OF COLON CANCER
Onrat ST1*, Çeken I2, Ellidokuz E3, Kupelioğlu A4
*Corresponding Author: Serap Tutgun Onrat, Department of Medical Genetics, Afyon Kocatepe University
Medical Faculty, ANS Arastırma Uygulama Hastanesi, Morfoloji Binası, Ozdilek yolu, Afyonkarahisar, 03200,
Turkey; Tel.: +90-272-246-3301, Fax: +90-272-246-3300, E-mail: tutgunonrat@ yahoo.com,
sonrat@aku.edu.tr
page: 25
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Abstract
Genetic alterations and changes in genomic DNA
cytosine methylation patterns are associated with all
types of cancer and are caused by germline mutations
in DNA mismatch repair (MMR) genes, predominantly
MLH1 (MutL homolog 1, 19 exons) and MSH2 (MutS
homolog 2, 16 exons). Genomic DNA was extracted
from tissue samples embedded in paraffin from 49 patients
with adenocarcinoma and from 21 patients with
carcinoma for the study group; genomic DNA was extracted
from lymphocytes from 10 healthy donors for
the control group. We used methylation specific multiplex
ligation-dependent probe amplification (MS-MLPA),
which allows the detection of copy number changes
and unusual methylation levels of 10 to 50 different
sequences in one reaction by use of the methylationsensitive
restriction enzyme HhaI and sequence-specific
capillary electrophoresis for the study of 24 genes.
We found the mean methylation rates for MLH1
(97.14%), MSH2 (24.28%), MSH6 (MutS homolog 6)
(67.14%), MSH3 (MutS homolog 3) (78.57%), MLH3
(MutL homolog 3) (75.71%), PMS2 (postmeiotic segregation
increased 2) (65.71%), MGMT(O-6-methylguanine-
DNA methyltransferase ) (82.85%). We conclude
that the mismatch repair (MMR) system is critical
for the maintenance of genomic stability.
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